Genotype-phenotype correlations in spastic paraplegia type 7: a study in a large Dutch cohort.
Spastic paraplegia type 7 is an autosomal recessive neurodegenerative disorder mainly characterized by progressive bilateral lower limb spasticity and referred to as a form of hereditary spastic paraplegia. Additional disease features may also be observed as part of a more complex phenotype. Many different mutations have already been identified, but no genotype-phenotype correlations have been found so far. From a total of almost 800 patients referred for testing, we identified 60 patients with mutations in the SPG7 gene. We identified 14 previously unreported mutations and detected a high recurrence rate of several earlier reported mutations. We were able to collect detailed clinical data for 49 patients, who were ranked based on a pure versus complex phenotype, ataxia versus no ataxia and missense versus null mutations. A generally complex phenotype occurred in 69% of all patients and was associated with a younger age at onset (trend with P = 0.07). Ataxia was observed in 57% of all patients. We found that null mutations were associated with the co-occurrence of cerebellar ataxia (trend with P = 0.06). The c.1409 G > A (p.Arg470Gln) mutation, which was found homozygously in two sibs, was associated with a specific complex phenotype that included predominant visual loss due to optical nerve atrophy. Neuropathology in one of these cases showed severe degeneration of the optic system, with less severe degeneration of the ascending tracts of the spinal cord and cerebellum. Other disease features encountered in this cohort included cervical dystonia, vertical gaze palsy, ptosis and severe intellectual disability. In this large Dutch cohort, we seem to have identified the first genotype-phenotype correlation in spastic paraplegia type 7 by observing an association between the cerebellar phenotype of spastic paraplegia type 7 and SPG7 null alleles. An overlapping phenotypic presentation with its biological counterpart AFG3L2, which when mutated causes spinocerebellar ataxia type 28, is apparent and possibly suggests that abnormal levels of the SPG7 protein impact the function of the mitochondrial ATPases associated with diverse cellular activities-protease complex (formed by SPG7 and AFG3L2) in the cerebellum. In addition, a missense mutation in exon 10 resulted in predominant optical nerve atrophy, which might suggest deleterious interactions of this SPG7 variant with its substrate OPA1, the mutated gene product in optic atrophy type 1. Functional studies are required to further investigate these interactions.


Measurement of glomerular filtration rate using dynamic magnetic resonance imaging in patients with chronic kidney disease.
Determination of glomerular filtration rate (GFR) using plasma disappearance curves requires the injection of a filtration marker and repeated timed blood collections. Gadolinium-containing contrast media are excreted exclusively by glomerular filtration and could provide a novel approach to quantifying GFR using magnetic resonance (MR) imaging. The aim of this study was to demonstrate the feasibility of measuring GFR by the clearance of gadolinium-containing contrast medium in patients with chronic kidney disease (CKD). Informed consent was obtained from stable CKD patients in stages 1, 2 or 3 (n=16; 5 women, 11 men; median age 54 years). GFR was measured after a bolus injection of gadobutrol (4 mL, approximately 0.05 mmol/kg) and calculated from the washout of the signal intensity obtained over the liver. The obtained MR-GFR was compared with simultaneously measured plasma clearance of inulin and gadobutrol. Technical failure occurred in 2 patients. The mean obtained MR-GFR was 71 ± 25 (SD) mL/min per 1.73 m² and agreed well with the mean inulin-GFR (70 ± 24 mL/min per 1.73 m²). Pearson's correlation coefficient was r=0.91. The mean of the paired differences was 1 ± 10 mL/min per 1.73 m² and not significantly different from zero. GFR obtained from gadobutrol plasma clearance also agreed well with inulin-GFR and MR-GFR (r=0.92 and r=0.75, respectively). We describe a novel method of determining GFR from MR imaging using a low dose of gadobutrol in patients with reduced GFR that enables the absolute quantification of GFR after routine contrast-enhanced MR imaging.


TB vaccines at the turn of the century: insights into immunity to M. tuberculosis and modern approaches for prevention of an ancient disease.
Tuberculosis (TB) remains a leading cause of infectious disease mortality worldwide, accounting for nearly 2 million deaths annually. Despite the availability of effective anti-TB therapy, the world's case burden of TB continues to climb, in part owing to the concurrent acquired immune deficiency syndrome pandemic. The widespread use of the current TB vaccine, the bacille Calmette-Guérin strain of M. bovis (BCG), has failed to curtail the TB epidemic and therefore strategies for the eradication of TB have centered on aggressive case-finding and managed treatment, such as directly observed treatment short course (DOTS). Although DOTS continues to have a significant impact, the ability of DOTS to eradicate TB is limited. Therefore, TB eradication will require the development of an improved vaccine, which, in turn, will require application of state-of-the-art vaccine technology and of our knowledge of TB immunology and TB genomics to this problem. In this article, knowledge about the requisite components of protective immunity, modern vaccine strategies, and elements of clinical trials required to evaluate vaccine efficacy, as well as the status of current TB vaccine candidates, are reviewed.


Featured Article: Nuclear export of opioid growth factor receptor is CRM1 dependent.
Opioid growth factor receptor (OGFr) facilitates growth inhibition in the presence of its specific ligand opioid growth factor (OGF), chemically termed [Met(5)]-enkephalin. The function of the OGF-OGFr axis requires the receptor to translocate to the nucleus. However, the mechanism of nuclear export of OGFr is unknown. In this study, endogenous OGFr, as well as exogenously expressed OGFr-EGFP, demonstrated significant nuclear accumulation in response to leptomycin B (LMB), an inhibitor of CRM1-dependent nuclear export, suggesting that OGFr is exported in a CRM1-dependent manner. One consensus sequence for a nuclear export signal (NES) was identified. Mutation of the associated leucines, L217 L220 L223 and L225, to alanine resulted in decreased nuclear accumulation. NES-EGFP responded to LMB, indicating that this sequence is capable of functioning as an export signal in isolation. To determine why the sequence functions differently in isolation than as a full length protein, the localization of subNES was evaluated in the presence and absence of MG132, a potent inhibitor of proteosomal degradation. MG132 had no effect of subNES localization. The role of tandem repeats located at the C-terminus of OGFr was examined for their role in nuclear trafficking. Six of seven tandem repeats were removed to form deltaTR. DeltaTR localized exclusively to the nucleus indicating that the tandem repeats may contribute to the localization of the receptor. Similar to the loss of cellular proliferation activity (i.e. inhibition) recorded with subNES, deltaTR also demonstrated a significant loss of inhibitory activity indicating that the repeats may be integral to receptor function. These experiments reveal that OGFr contains one functional NES, L217 L220 L223 and L225 and can be exported from the nucleus in a CRM1-dependent manner.


Highly selective tridentate fluorescent probes for visualizing intracellular Mg2+ dynamics without interference from Ca2+ fluctuation.
We developed highly selective fluorescent Mg2+ probes based on a novel tridentate Mg2+-selective chelator. The superior selectivity for Mg2+ over Ca2+ enabled the detection of intracellular Mg2+ concentration changes without any response to Ca2+ influx. The new probes are potential keys to elucidating intracellular Mg2+ dynamics under conditions of extremely sharp Ca2+ fluctuation.


Working at the nexus between public health policy, practice and research. Dynamics of knowledge sharing in The Netherlands.
Joining the domains of practice, research and policy is an important aspect of boosting the quality performance required to tackle complex public health problems. "Joining domains" implies a departure from the linear and technocratic knowledge-translation approach. Integrating the practice, research and policy triangle means knowing its elements, appreciating the barriers, identifying possible cooperation strategies and studying strategy effectiveness under specified conditions.This article examines the dynamic process of developing an Academic Collaborative Centre for Public Health in the Netherlands, with the objective of achieving that the three domains of policy, practice and research become working partners on an equal footing. An interpretative hermeneutic approach was used to interpret the phenomenon of collaboration at the nexus between the three domains. The project was explicitly grounded in current organizational culture and routines, applied to nexus action. In the process of examination, we used both quantitative (e.g. records) and qualitative data (e.g., interviews and observations). The data were interpreted using the Actor-Network, Institutional Re-Design and Blurring the Boundaries theories. Results show commitment at strategic level. At the tactical level, however, managers were inclined to prioritize daily routine, while the policy domain remained absent. At the operational level, practitioners learned to do PhD research in real-life practice and researchers became acquainted with problems of practice and policy, resulting in new research initiatives. We conclude that working at the nexus is an ongoing process of formation and reformation. Strategies based on Institutional Re-Design theories in particular might help to more actively stimulate managers' involvement to establish mutually supportive networks.


The Effectiveness of Mindfulness- and Acceptance-Based Interventions for Informal Caregivers of People With Dementia: A Meta-Analysis.
The application of mindfulness- and acceptance-based interventions (MABIs) for informal caregivers of people with dementia (PwD) is relatively novel, and the current state of the evidence base is unclear. This meta-analysis examined the effectiveness of MABIs on reducing symptoms of depression and burden in informal caregivers of PwD. The quality of included studies was evaluated and moderator variables explored. A literature search of six electronic databases (PsycARTICLES, PsycINFO, MEDLINE Complete, SCOPUS, Web of Science, and ProQuest) was conducted from the first available date to 20 December 2016. Inclusion criteria involved studies that quantitatively investigated the impact of MABIs on depression and/or burden in informal caregivers of PwD. Twelve studies, providing data on 321 caregivers, were included. Most used mindfulness-based stress reduction and were conducted in the United States. The average attrition among participants was 15.83%. The pre-post effect of MABIs was large for depression and moderate for burden. These effects were largely maintained at follow-up. Significant heterogeneity of effect sizes was observed, with no significant moderators identified. Study quality varied from very poor to moderately good. The low attrition and moderate to large effects suggest that MABIs are acceptable and beneficial for informal caregivers of PwD. The lack of significant moderators could advocate services using more cost-effective forms of MABIs. Further higher-quality research is needed to improve the robustness of the evidence base and enable a meta-analysis to thoroughly examine and quantify moderator variables.


Non-invasive prediction of reperfusion and coronary artery patency by continuous ST segment monitoring in the GUSTO-I trial.
In the GUSTO-I ECG ischaemia monitoring substudy, 1067 patients underwent continuous ST segment monitoring, using vector-derived 12-lead (406 patients), 12-lead (373 patients) and 3-lead Holter (288 patients) ECG recording systems. Simultaneous angiograms at 90 or 180 min following thrombolytic therapy were performed as a part of the prospective study in 302 patients. Infarct vessel patency was established as TIMI perfusion grades 2 or 3 and occlusion as TIMI perfusion grades 0 or 1. Coronary artery patency was predicted from ST trends up to the time of angiography. Predictive values at 90 and 180 min after the start of thrombolysis were 70% and 82% for patency and 58% and 64% for occlusion, respectively. In retrospect, accuracy appeared greatest (79-100%) in patients with extensive ST segment elevation (> or = 400 microV), if both speed of ST recovery and extent of ST segment elevation were taken into account. Although the three recording systems differed considerably in signal processing, no significant difference in accuracy was demonstrated among these systems. We conclude that continuous ECG monitoring may help select high risk patients without apparent reperfusion who may benefit from additional reperfusion therapy. As ST recovery may occur early after the start of thrombolytics and accuracy of the test is related to peak ST levels, the use of on-line ECG monitoring devices on emergency wards and cardiac care units is recommended.


Standardized interpretation of paediatric chest radiographs for the diagnosis of pneumonia in epidemiological studies.
Although radiological pneumonia is used as an outcome measure in epidemiological studies, there is considerable variability in the interpretation of chest radiographs. A standardized method for identifying radiological pneumonia would facilitate comparison of the results of vaccine trials and epidemiological studies of pneumonia. A WHO working group developed definitions for radiological pneumonia. Inter-observer variability in categorizing a set of 222 chest radiographic images was measured by comparing the readings made by 20 radiologists and clinicians with a reference reading. Intra-observer variability was measured by comparing the initial readings of a randomly chosen subset of 100 radiographs with repeat readings made 8-30 days later. Of the 222 images, 208 were considered interpretable. The reference reading categorized 43% of these images as showing alveolar consolidation or pleural effusion (primary end-point pneumonia); the proportion thus categorized by each of the 20 readers ranged from 8% to 61%. Using the reference reading as the gold standard, 14 of the 20 readers had sensitivity and specificity of > 0.70 in identifying primary end-point pneumonia; 13 out of 20 readers had a kappa index of > 0.6 compared with the reference reading. For the 92 radiographs deemed to be interpretable among the 100 images used for intra-observer variability, 19 out of 20 readers had a kappa index of > 0.6. Using standardized definitions and training, it is possible to achieve agreement in identifying radiological pneumonia, thus facilitating the comparison of results of epidemiological studies that use radiological pneumonia as an outcome.


Structural Transitions in Ceramide Cubic Phases during Formation of the Human Skin Barrier.
The stratum corneum is the outermost layer of human skin and the primary barrier toward the environment. The barrier function is maintained by stacked layers of saturated long-chain ceramides, free fatty acids, and cholesterol. This structure is formed through a reorganization of glycosylceramide-based bilayers with cubic-like symmetry into ceramide-based bilayers with stacked lamellar symmetry. The process is accompanied by deglycosylation of glycosylceramides and dehydration of the skin barrier lipid structure. Using coarse-grained molecular dynamics simulation, we show the effects of deglycosylation and dehydration on bilayers of human skin glycosylceramides and ceramides, folded in three dimensions with cubic (gyroid) symmetry. Deglycosylation of glycosylceramides destabilizes the cubic lipid bilayer phase and triggers a cubic-to-lamellar phase transition. Furthermore, subsequent dehydration of the deglycosylated lamellar ceramide system closes the remaining pores between adjacent lipid layers and locally induces a ceramide chain transformation from a hairpin-like to a splayed conformation.


Identification of a Streptococcus gordonii SspB domain that mediates adhesion to Porphyromonas gingivalis.
Porphyromonas gingivalis, a primary pathogen in adult periodontitis, may establish itself in the oral cavity by adhering to early plaque bacteria such as Streptococcus gordonii. Our previous studies (R. J. Lamont et al., Microbiology 140:867-872, 1994) suggested that this interaction is mediated by the SspB polypeptide, a member of the antigen I/II family of streptococcal surface proteins. S. gordonii was recently shown to express a second Ssp polypeptide (SspA) that resembles SspB and the structurally homologous antigen I/II polypeptide (Pac) of Streptococcus mutans. To determine if all of these related antigen I/II proteins interacted with P. gingivalis, SspA, SspB, and Pac were tested for adhesion to P. gingivalis cells. Both of the S. gordonii Ssp proteins bound labeled target cells, whereas the S. mutans Pac polypeptide did not, suggesting that antigen I/II-mediated binding of P. gingivalis by streptococci may be species specific. To investigate the molecular basis for this functional difference, the P. gingivalis binding domain of SspB was mapped. The binding properties of a family of truncated SspB polypeptides lacking C-terminal sequences were determined. In addition, the lack of binding activity exhibited by the Pac protein was exploited to construct and analyze chimeric SspB-Pac polypeptides. Both approaches revealed that the region defined by residues 1167 to 1250 of SspB was essential for P. gingivalis binding. This region of SspA and SspB is entirely conserved, consistent with the binding properties determined for these proteins. However, the corresponding region of Pac differs in both the primary sequence and predicted secondary structure, suggesting that the overall structure of this domain may define its functional activity.


The assessment of biases in the acoustic discrimination of individuals.
Animal vocalizations contain information about individual identity that could potentially be used for the monitoring of individuals. However, the performance of individual discrimination is subjected to many biases depending on factors such as the amount of identity information, or methods used. These factors need to be taken into account when comparing results of different studies or selecting the most cost-effective solution for a particular species. In this study, we evaluate several biases associated with the discrimination of individuals. On a large sample of little owl male individuals, we assess how discrimination performance changes with methods of call description, an increasing number of individuals, and number of calls per male. Also, we test whether the discrimination performance within the whole population can be reliably estimated from a subsample of individuals in a pre-screening study. Assessment of discrimination performance at the level of the individual and at the level of call led to different conclusions. Hence, studies interested in individual discrimination should optimize methods at the level of individuals. The description of calls by their frequency modulation leads to the best discrimination performance. In agreement with our expectations, discrimination performance decreased with population size. Increasing the number of calls per individual linearly increased the discrimination of individuals (but not the discrimination of calls), likely because it allows distinction between individuals with very similar calls. The available pre-screening index does not allow precise estimation of the population size that could be reliably monitored. Overall, projects applying acoustic monitoring at the individual level in population need to consider limitations regarding the population size that can be reliably monitored and fine-tune their methods according to their needs and limitations.


Understanding methods for estimating HIV-associated maternal mortality.
The impact of HIV on maternal mortality and more broadly on the health of women, remains poorly documented and understood. Two recent reports attempt to address the conceptual and methodological challenges that arise in estimating HIV-related maternal mortality and trends. This paper presents and compares the methods and discusses how they affect estimates at global and regional levels. Country examples of likely patterns of mortality among women of reproductive age are provided to illustrate the critical interactions between HIV and complications of pregnancy in high-HIV-burden countries. The implications for collaboration between HIV and reproductive health programmes are discussed, in support of accelerated action to reach the Millennium Development Goals and improve the health of women.


Increase in oxidative stress as measured by cerebrospinal fluid lipid peroxidation during treatment for childhood acute lymphoblastic leukemia.
Five-year survival from childhood acute lymphoblastic leukemia (ALL) approaches 90%, but 40% of survivors experience central nervous system (CNS) treatment-related cognitive problems. Despite considerable evidence for cognitive problems, less is known about mechanisms of neurological injury. Our purpose was to investigate oxidative stress, measured by lipid peroxidation, as a mechanism of CNS treatment-related neurological injury. The sample included 55 children (mean age at diagnosis=6.84 y, SD=3.40) who received intrathecal and intravenous chemotherapy for CNS-directed treatment according to Children's Oncology Group protocols. Glycerophospholipids were extracted from cerebrospinal fluid samples obtained at diagnosis and during intrathecal chemotherapy administration. Unoxidized and oxidized phosphatidylcholine (PC) and phosphatidylinositol (PI) were measured by normal phase high-performance liquid chromatography with diode array detection, and analyzed with a general linear model for repeated measures analysis of variance. Compared with the diagnostic cerebrospinal fluid sample, unoxidized and oxidized PC and PI increased significantly across treatment phases. Amount of intravenous methotrexate received was significantly correlated with oxidized PI, and age at time of ALL diagnosis was significantly associated with oxidized PC. These findings support our hypothesis that oxidative stress is a mechanism of neurological injury associated with CNS-directed treatment for ALL.


Infrared Laser Activation of Soluble and Membrane Protein Assemblies in the Gas Phase.
Collision-induced dissociation (CID) is the dominant method for probing intact macromolecular complexes in the gas phase by means of mass spectrometry (MS). The energy obtained from collisional activation is dependent on the charge state of the ion and the pressures and potentials within the instrument: these factors limit CID capability. Activation by infrared (IR) laser radiation offers an attractive alternative as the radiation energy absorbed by the ions is charge-state-independent and the intensity and time scale of activation is controlled by a laser source external to the mass spectrometer. Here we implement and apply IR activation, in different irradiation regimes, to study both soluble and membrane protein assemblies. We show that IR activation using high-intensity pulsed lasers is faster than collisional and radiative cooling and requires much lower energy than continuous IR irradiation. We demonstrate that IR activation is an effective means for studying membrane protein assemblies, and liberate an intact V-type ATPase complex from detergent micelles, a result that cannot be achieved by means of CID using standard collision energies. Notably, we find that IR activation can be sufficiently soft to retain specific lipids bound to the complex. We further demonstrate that, by applying a combination of collisional activation, mass selection, and IR activation of the liberated complex, we can elucidate subunit stoichiometry and the masses of specifically bound lipids in a single MS experiment.


Design, synthesis, and X-ray analysis of a glycoconjugate bound to Mycobacterium tuberculosis antigen 85C.
Tuberculosis (TB) is a global health threat with nearly 500 000 new cases of multidrug-resistant TB estimated to occur every year, so new drugs are desperately needed. A number of current antimycobacterial drugs work by interfering with the biosynthesis of key components of the mycolylarabinogalactan (mAG). In light of this observation, other enzymes involved in the synthesis of the mAG should also serve as targets for antimycobacterial drug development. One potential target is the Antigen 85 (Ag85) complex, a family of mycolyltransferases that are responsible for the transfer of mycolic acids from trehalose monomycolate (TMM) to the arabinogalactan. Virtual thiophenyl-arabinoside conjugates were docked to antigen Ag85C (PDB code: 1va5 ) using Glide. Compounds with good docking scores were synthesized by a Gewald synthesis followed by linking to 5-thioarabinofuranosides. The resulting thiophenyl-thioarabinofuranosides were assayed for inhibition of mycoyltransferase activity using a 4-methylumbelliferyl butyrate fluorescence assay. The conjugates showed K(i) values ranging from 18.2 to 71.0 μM. The most potent inhibitor was soaked into crystals of Mycobacterium tuberculosis antigen 85C and the structure of the complex determined. The X-ray structure shows the compound bound within the active site of the enzyme with the thiophene moiety positioned in the putative α-chain binding site of TMM and the arabinofuranoside moiety within the known carbohydrate-binding site as exhibited for the Ag85B-trehalose crystal structure. Unexpectedly, no specific hydrogen bonding interactions are being formed between the arabinofuranoside and the carbohydrate-binding site of the active site suggesting that the binding of the arabinoside within this structure is driven by shape complementarily between the arabinosyl moiety and the carbohydrate binding site.


Minocycline protects photoreceptors from light and oxidative stress in primary bovine retinal cell culture.
To determine whether minocycline, a compound known to protect the retina against light-induced damage in rodent models, and its structurally related analogues would protect photoreceptor cells in primary bovine retinal cell culture against light and oxidative stress. Minocycline and its analogues were tested in primary retinal cell culture to see whether they would inhibit light or oxidative stress-induced cell death. Primary cell cultures composed of photoreceptors, bipolar cells, and glial cells were prepared from bovine retinas. The extent of cell death induced by light or oxidative stress was assessed by using Sytox Green (Invitrogen-Molecular Probes, Eugene, OR) a nucleic acid dye uptake assay. Differential protection of photoreceptor cells from stress were examined using immunocytochemistry. Minocycline and methacycline were cytoprotective against light- or oxidative stress-induced damage of bovine primary photoreceptors in culture with an EC(50) < 10 microM. In contrast, structurally related analogues such as demeclocycline, meclocycline, and doxycycline were phototoxic at >3 to >10 microM. Though demeclocycline was found to be phototoxic, it was cytoprotective (EC(50) = 5 microM) against oxidative stress in the absence of exposure to light. The protective action of minocycline against light-induced damage in the cell-based assays agrees with earlier reports in animal models and suggests that the in vitro assay using bovine primary retinal cell culture is a suitable model for evaluating compounds for retinal protection. Cellular protection or toxicity produced by structurally related compounds show that minor structural modifications can alter the function of minocycline and lead to potent retinal protective compounds.


Unplanned readmission rates, length of hospital stay, mortality, and medical costs of ten common medical conditions: a retrospective analysis of Hong Kong hospital data.
Studies on readmissions attributed to particular medical conditions, especially heart failure, have generally not addressed the factors associated with readmissions and the implications for health outcomes and costs. This study aimed to investigate the factors associated with 30-day unplanned readmission for 10 common conditions and to determine the cost implications. This population-based retrospective cohort study included patients admitted to all public hospitals in Hong Kong in 2007. The sample consisted of 337,694 hospitalizations in Internal Medicine. The disease-specific risk-adjusted odd ratio (OR), length of stay (LOS), mortality and attributable medical costs for the year were examined for unplanned readmissions for 10 medical conditions, namely malignant neoplasms, heart diseases, cerebrovascular diseases, pneumonia, injury and poisoning, nephritis and nephrosis, diabetes mellitus, chronic liver disease and cirrhosis, septicaemia, and aortic aneurysm. The overall unplanned readmission rate was 16.7%. Chronic liver disease and cirrhosis had the highest OR (1.62, 95% confidence interval (CI) 1.39-1.87). Patients with cerebrovascular disease had the longest LOS, with mean acute and rehabilitation stays of 6.9 and 3.0 days, respectively. Malignant neoplasms had the highest mortality rate (30.8%) followed by aortic aneurysm and pneumonia. The attributed medical cost of readmission was highest for heart disease (US$3 199 418, 95% CI US$2 579 443-803 393). Our findings showed variations in readmission rates and mortality for different medical conditions which may suggest differences in the quality of care provided for various medical conditions. In-hospital care, comprehensive discharge planning, and post-discharge community support for patients need to be reviewed to improve the quality of care and patient health outcomes.


Elucidation of the Erwinia uredovora carotenoid biosynthetic pathway by functional analysis of gene products expressed in Escherichia coli.
The most important function of carotenoid pigments, especially beta-carotene in higher plants, is to protect organisms against photooxidative damage (G. Britton, in T. W. Goodwin, ed., Plant Pigments--1988, 1988; N. I. Krinsky, in O. Isler, H. Gutmann, and U. Solms, ed., Carotenoids--1971, 1971). beta-Carotene also functions as a precursor of vitamin A in mammals (G. A. J. Pitt, in I. Osler, H. Gutmann, and U. Solms, ed., Carotenoids--1971, 1971). The enzymes and genes which mediate the biosynthesis of cyclic carotenoids such as beta-carotene are virtually unknown. We have elucidated for the first time the pathway for biosynthesis of these carotenoids at the level of enzyme-catalyzed reactions, using bacterial carotenoid biosynthesis genes. These genes were cloned from a phytopathogenic bacterium, Erwinia uredovora 20D3 (ATCC 19321), in Escherichia coli and located on a 6,918-bp fragment whose nucleotide sequence was determined. Six open reading frames were found and designated the crtE, crtX, crtY, crtI, crtB, and crtZ genes in reference to the carotenoid biosynthesis genes of a photosynthetic bacterium, Rhodobacter capsulatus; only crtZ had the opposite orientation from the others. The carotenoid biosynthetic pathway in Erwinia uredovora was clarified by analyzing carotenoids accumulated in E. coli transformants in which some of these six genes were expressed, as follows: geranylgeranyl PPiCrtB----prephytoene PPiCrtE----phytoeneCrtI---- lycopeneCrtY----beta-caroteneCrtZ----zeaxanthinCrtX--- -zeaxanthin-beta- diglucoside. The carotenoids in this pathway appear to be close to those in higher plants rather than to those in bacteria. Also significant is that only one gene product (CrtI) for the conversion of phytoene to lycopene is required, a conversion in which four sequential desaturations should occur via the intermediates phytofluene, zeta-carotene, and neurosporene.


Tea and cancer prevention: molecular mechanisms and human relevance.
Tea made from the leaves of the plant Camellia sinensis is a popular beverage. The possible cancer-preventive activity of tea and tea polyphenols has been studied extensively. This article briefly reviews studies in animal models, cell lines, and possible relevance of these studies to the prevention of human cancer. The cancer-preventive activity of tea constituents have been demonstrated in many animal models including cancer of the skin, lung, oral cavity, esophagus, stomach, liver, pancreas, small intestine, colon, bladder, prostate, and mammary gland. The major active constituents are polyphenols, of which (-)-epigallocatechin-3-gallate (EGCG) is most abundant, most active, and most studied, and caffeine. The molecular mechanisms of the cancer-preventive action, however, are just beginning to be understood. Studies in cell lines led to the proposal of many mechanisms on the action of EGCG. However, mechanisms based on studies with very high concentrations of EGCG may not be relevant to cancer prevention in vivo. The autooxidation of EGCG in cell culture may also produce activities that do not occur in many internal organs. In contrast to the cancer prevention activity demonstrated in different animal models, no such conclusion can be convincingly drawn from epidemiological studies on tea consumption and human cancers. Even though the human data are inconclusive, tea constituents may still be used for the prevention of cancer at selected organ sites if sufficient concentrations of the agent can be delivered to these organs. Some interesting examples in this area are discussed.


Identification, Evaluation, and Management of Children With Autism Spectrum Disorder.
Autism spectrum disorder (ASD) is a common neurodevelopmental disorder with reported prevalence in the United States of 1 in 59 children (approximately 1.7%). Core deficits are identified in 2 domains: social communication/interaction and restrictive, repetitive patterns of behavior. Children and youth with ASD have service needs in behavioral, educational, health, leisure, family support, and other areas. Standardized screening for ASD at 18 and 24 months of age with ongoing developmental surveillance continues to be recommended in primary care (although it may be performed in other settings), because ASD is common, can be diagnosed as young as 18 months of age, and has evidenced-based interventions that may improve function. More accurate and culturally sensitive screening approaches are needed. Primary care providers should be familiar with the diagnostic criteria for ASD, appropriate etiologic evaluation, and co-occurring medical and behavioral conditions (such as disorders of sleep and feeding, gastrointestinal tract symptoms, obesity, seizures, attention-deficit/hyperactivity disorder, anxiety, and wandering) that affect the child's function and quality of life. There is an increasing evidence base to support behavioral and other interventions to address specific skills and symptoms. Shared decision making calls for collaboration with families in evaluation and choice of interventions. This single clinical report updates the 2007 American Academy of Pediatrics clinical reports on the evaluation and treatment of ASD in one publication with an online table of contents and section view available through the American Academy of Pediatrics Gateway to help the reader identify topic areas within the report.


Synthesis and preliminary anti-inflammatory and anti-bacterial evaluation of some diflunisal aza-analogs.
Our aim was to identify new multi-target compounds endowed with both anti-inflammatory and anti-bacterial activities for treatment of human infections. Diflunisal, a nonsteroidal anti-inflammatory agent, has recently been repurposed for its anti-virulence properties against methicillin-resistant Staphylococcus aureus. Effective synthesis of some aza-analogs of the anti-inflammatory drug diflunisal was carried out following the route involving key oxazole intermediates to obtain o- and m-hydroxypyridinecarboxylic acid derivatives. The newly synthesized diflunisal aza-analogs did not exhibit cytotoxic activity up to 80 μM and some of them exhibited anti-inflammatory activities, decreasing the levels of pro-inflammatory cytokines and prostaglandins induced by bacterial lipopolysaccharide in human primary macrophages. Ten of the diflunisal aza-analogs were found to have interesting antibacterial activity, sensitizing S. aureus, Streptococcus pyogenes, Enterococcus faecium, and Pseudomonas aeruginosa to the antibacterial effects of beta-lactam antibiotics and protein synthesis inhibitors.


Performance of a commercial rapid scanning spectrometer in emission and absorption.
The performance of a commercial rapid scanning spectrometer was checked experimentally in studies of flash photolysis, photochromism, phosphorescence, and thermal radiation. Rapid scan spectra (1 msec to 100 msec) were measured in portions of the uv, visible, and ir spectral regions (total wavelength coverage 250 nm to 14.5 microm), at rates up to 800 spectra/sec. The effect of grating line spacing on resolution and spectral range was measured. In absorption measurements, successive scans of the background radiation source were photometrically reproducible to within 1 part in 80. The effect of scanning time on resolution was studied experimentally; resolution of 0.94 cm(-1) at 2400 cm(-1) was achieved with signal-to-noise of 30:1, during rapid scanning. Recording on magnetic tape and oscillographic recording are illustrated.


An amino acid outside the pore region influences apamin sensitivity in small conductance Ca2+-activated K+ channels.
Small conductance calcium-activated potassium channels (SK, K(Ca)) are a family of voltage-independent K+ channels with a distinct physiology and pharmacology. The bee venom toxin apamin inhibits exclusively the three cloned SK channel subtypes (SK1, SK2, and SK3) with different affinity, highest for SK2, lowest for SK1, and intermediate for SK3 channels. The high selectivity of apamin made it a valuable tool to study the molecular makeup and function of native SK channels. Three amino acids located in the outer vestibule of the pore are of particular importance for the different apamin sensitivities of SK channels. Chimeric SK1 channels, enabling the homomeric expression of the rat SK1 (rSK1) subunit and containing the core domain (S1-S6) of rSK1, are apamin-insensitive. By contrast, channels formed by the human orthologue human SK1 (hSK1) are sensitive to apamin. This finding hinted at the involvement of regions beyond the pore as determinants of apamin sensitivity, because hSK1 and rSK1 have an identical amino acid sequence in the pore region. Here we investigated which parts of the channels outside the pore region are important for apamin sensitivity by constructing chimeras between apamin-insensitive and -sensitive SK channel subunits and by introducing point mutations. We demonstrate that a single amino acid situated in the extracellular loop between the transmembrane segments S3 and S4 has a major impact on apamin sensitivity. Our findings enabled us to convert the hSK1 channel into a channel that was as sensitive for apamin as SK2, the SK channel with the highest sensitivity.


Clinical Frailty Scale: Translation and Cultural Adaptation into the Brazilian Portuguese Language.
Frailty is a biological syndrome that causes adverse events in the health of older adults. However, the Clinical Frailty Scale has not yet been culturally adapted and validated into Brazilian Portuguese language. Our aim was to translate, reproduce and validate the Clinical Frailty Scale (CFS) for the Brazilian Portuguese language. An observational cross-sectional study with senior patients was conducted between Jan 2018 and Nov 2018. Volunteers aged >60 and living in Brazil. The translation and cultural adaptation of the CFS into the Portuguese language, the principles and good practices were followed. To conduct the validation and determine the reproducibility of an inter-observer evaluation, the patients answered the scale questions in Portuguese on two occasions, delivered by two separate examiners and separated by a 10-minute interval, on their first visit; the 36-item Short Form Survey quality-of-life questionnaire (SF-36) was also applied. Seven days later, a second visit was undertaken to perform an intra-observer reproducibility assessment. A total of 66 older individuals were enrolled (72 ± 8 years), the majority of which did not present frailty (63.6%) and reported a low physical limitation level in the SF-36. The CFS showed a significant correlation with the SF-36 quality-of-life questionnaire (r= -0.663; p<0.0001) and no statistical difference was observed between intra-rater (p=0.641) and inter-rater (p=0.350) applications, demonstrating the reproducibility and applicability of the instrument. The standard error estimate (SEE) was evaluated and there were no differences between the CFS and the SF-36 (SEE= 1.13 points). The Brazilian Portuguese language version of the CFS is a valid, reproducible and reliable instrument for evaluating the impact of frailty on the lives of senior patients.


A knowledge translation tool improved osteoporosis disease management in primary care: an interrupted time series analysis.
Osteoporosis affects over 200 million people worldwide at a high cost to healthcare systems, yet gaps in management still exist. In response, we developed a multi-component osteoporosis knowledge translation (Op-KT) tool involving a patient-initiated risk assessment questionnaire (RAQ), which generates individualized best practice recommendations for physicians and customized education for patients at the point of care. The objective of this study was to evaluate the effectiveness of the Op-KT tool for appropriate disease management by physicians. The Op-KT tool was evaluated using an interrupted time series design. This involved multiple assessments of the outcomes 12 months before (baseline) and 12 months after tool implementation (52 data points in total). Inclusion criteria were family physicians and their patients at risk for osteoporosis (women aged ≥ 50 years, men aged ≥ 65 years). Primary outcomes were the initiation of appropriate osteoporosis screening and treatment. Analyses included segmented linear regression modeling and analysis of variance. The Op-KT tool was implemented in three family practices in Ontario, Canada representing 5 family physicians with 2840 age eligible patients (mean age 67 years; 76% women). Time series regression models showed an overall increase from baseline in the initiation of screening (3.4%; P < 0.001), any osteoporosis medications (0.5%; P = 0.006), and calcium or vitamin D (1.2%; P = 0.001). Improvements were also observed at site level for all the three sites considered, but these results varied across the sites. Of 351 patients who completed the RAQ unprompted (mean age 64 years, 77% women), the mean time for completing the RAQ was 3.43 minutes, and 56% had any disease management addressed by their physician. Study limitations included the inherent susceptibility of our design compared with a randomized trial. The multicomponent Op-KT tool significantly increased osteoporosis investigations in three family practices, and highlights its potential to facilitate patient self-management. Next steps include wider implementation and evaluation of the tool in primary care.


"They Destroy the Reproductive System": Exploring the Belief that Modern Contraceptive Use Causes Infertility.
A common reason for nonuse of modern contraceptives is concern about side effects and health complications. This article provides a detailed characterization of the belief that modern contraceptives cause infertility, and an examination of how this belief arises and spreads, and why it is so salient. We conducted focus group discussions and key informant interviews in three rural communities along Kenya's eastern coast, and identified the following themes: (1) the belief that using modern contraception at a young age or before childbirth can make women infertile is widespread; (2) according to this belief, the most commonly used methods in the community were linked to infertility; (3) when women observe other women who cannot get pregnant after using modern contraceptives, they attribute the infertility to the use of contraception; (4) within the communities, the primary goal of marriage is childbirth and thus community approval is rigidly tied to childbearing; and, therefore (5) the social consequences of infertility are devastating. These findings may help inform the design of programs to address this belief and reduce unmet need.


Sequential modulation of cue use in the task switching paradigm.
In task switching studies, pre-cuing of the upcoming task improves performance, indicating preparatory activation of the upcoming task-set, and/or inhibition of the previous task-set. To further investigate cue-based task preparation, the authors presented both valid and invalid task cues in a task switching experiment involving three tasks. Consistent with previous findings, a validity effect in terms of higher reaction times on invalidly compared to validly cued tasks was obtained. However, this validity effect was reduced following invalidly cued trials, suggesting dynamic adjustment in terms of decreased cue-based preparation after being misled. Performance was particularly impaired when the current task was the one that was invalidly cued on the preceding trial. This finding may reflect either particular reluctance to prepare or persisting inhibition of the erroneously prepared task-set from the pre-trial.


Immunomodulatory effect of procainamide in man. Inhibition of human suppressor T-cell activity in vitro.
Procainamide (PA) induces the production of a number of autoantibodies in a high proportion of treated individuals and in some a syndrome closely resembling systemic lupus erythematosus. The mechanism underlying this action of PA is unclear. To examine the possibility that PA might induce autoantibody formation by altering normal immunoregulatory mechanisms, the action of this drug on an in vitro model of antibody formation in man was examined. PA was found to augment the generation of immunoglobulin-secreting cells (ISC) from human peripheral blood mononuclear cells (PBM) in response to pokeweed mitogen but had no effect on pokeweed mitogen-induced tritiated thymidine incorporation. When purified populations of B and T cells were used, PA enhanced the generation of ISC in B-cell cultures supported by untreated T cells but not by T cells treated with mitomycin C. These results indicate that PA augmented B-cell responses by inhibiting suppressor T-cell activity and not by augmenting helper T-cell or B-cell function. N-Acetyl-procainamide had no effect on the generation of ISC in this system. The effect of PA on concanavalin A (Con A)-induced suppressor cell activity was also examined to determine whether PA altered the generation or expression of suppressor T-cell function. PBM were cultured with 30 microgram/ml of Con A for 48 h to generate suppressor cells. When these were co-cultured with fresh PBM, the number of ISC generated was decreased by 58.1 +/- 3.4% (mean +/- SEM, n = 6). Cells that had been similarly incubated without Con A were not inhibitory. The addition of PA to the Con A-stimulated cultures inhibited the generation of suppressor cells as indicated by the fact that the response of fresh cells co-cultured with the Con A-stimulated cells was diminished by only 27.2 +/- 4.3%. In this system too, N-acetyl-procaimamide had no effect. By contrast, adding PA only to the co-culture of Con A-stimulated cells with fresh PBM had a less marked effect on suppressor cell function. These results indicate that the major action of PA is to inhibit the generation of suppressor T-cell activity. Such an effect may explain the capacity of this agent to induce autoantibody formation in treated individuals.


Enhancement of low-contrast curvilinear features in imagery.
A new method is described for enhancing low-contrast curvilinear features in imagery that combines directional filtering with Fischler, Tenenbaum and Wolf's F* algorithm for computing minimum cost paths. The method exploits a phenomenon called "the stability of lines over angle." The idea is that when a directionally filtered image contains a line plus noise, minimum cost paths tend to be aligned in the direction of the line with random jumps between parallel paths. When the input image contains noise only, the direction of minimum cost paths resemble random walks with drift. As the direction of the filter changes, minimum cost paths that follow true features persist and are more stable over angle than those that follow noise. Adding them up in an accumulator array over angle produces a larger number of votes along signal paths than along noise paths. This provides a means for enhancing trajectories of low-contrast features. Several examples illustrate the enhancement of forest trails in USGS aerial imagery, linear features on Mars, and roads in synthetic aperture radar imagery.


Factors contributing to non-randomness in species Co-occurrences on Islands.
There has been dispute whether patterns of species co-occurrence on islands are largely random. We present a new method for testing this question; this method lets one not only examine whether a whole fauna is non-randomly structured, but also identify in which direction and by how much each particular species combination deviates from expectations based on randomness. Application of this method to the whole Bismarck and New Hebridean avifaunas, and to two particular guilds, shows that some pairs of species have more exclusive distributions than expected for random placement of species, because of competition, differing distributional strategies, or different geographical orgins. Other pairs of species have more coincident distributions than expected, because of shared habitat, single-island endemism, shared distributional strategies, or shared geographical origins. Much of the information about non-random co-occurrence is contained in the incidence graphs for occurrence of individual species. Finally, our present understanding of assembly rules is summarized.


Directed assembly of functionalized nanoparticles with amphiphilic diblock copolymers.
The ability to design and fabricate highly ordered superstructures from nanoscale particles remains a major scientific and technological challenge. Patchy nanoparticles have recently emerged as a novel class of building units to construct functional materials. Using simulations of coarse-grained molecular dynamics, we propose a simple approach to achieve soft nanoparticles with a self-patchiness nature through self-assembly of tethered copolymers with a sequence of inner solvophilic and outer solvophobic blocks. As building units, the patch-like nanoparticles are directed to further assemble into a rich variety of highly ordered superstructures via condensation-coalescence mechanisms. The growth kinetics of the superstructures obeys the kinetic model of the step-growth polymerization process. Our simulations also demonstrate that the intermediate patch-like nanoparticles and the final assembled superstructures can be rationally tuned by changing the number and the composition of the tethered copolymer chains. This strategy of copolymer functionalization conceptually enables the design and fabrication of highly ordered superstructures of nanoparticle ensembles with new horizons for promising applications in soft nanotechnology and biotechnology.


Evaluation of asthma control by physicians and patients: comparison with current guidelines.
Current asthma consensus guidelines recommend a series of criteria for determining whether asthma is controlled. It is not known whether physicians are using these criteria to assess treatment needs and how effective such assessments are compared with patient assessment of asthma control. To compare the parameters used by physicians and patients with asthma to determine whether asthma control is acceptable, according to the current Canadian asthma consensus guidelines. A total of 183 Canadian physicians, mostly general practitioners, evaluated 856 patients with mildly to moderately uncontrolled asthma who were not using anti-inflammatory medications at the time of entry in the study. Physician characteristics and patient demographics were obtained. The physicians completed two questionnaires, one assessing the level of asthma control of the patient on an ordinal scale from 1 (very poor) to 5 (very good) and another indicating the parameters that were used to evaluate this level of control. Patients answered an asthma control questionnaire identical to the one completed by the physician and completed a six-question asthma control questionnaire, with each question scored on a 0- to 6-point scale. Although according to current asthma guidelines all patients surveyed had uncontrolled asthma, 66.2% of patients and 43.3% of physicians rated control of asthma symptoms as adequate to very good. The average scores for patient- and physician-rated asthma control were 3.0 0.2 and 2.6 0.2, respectively. The average patient score on the Juniper asthma questionnaire was 12.2 6.3. Physicians used a mean of seven parameters to assess the patient's level of asthma control, mostly beta2-agonist need, followed by cough, wheezing, shortness of breath, limitation of physical activities and night-time awakenings. Pediatricians used cough more frequently as an evaluation parameter, and respirologists measured pulmonary function more often than other physcians. Some parameters not usually included in guideline criteria for control, such as fatigue, need to clear throat, colored sputum, headache and dizziness, were sometimes used by physicians. Only 10% and 18% of physicians used measurements of forced expiratory volume in 1 s and peak expiratory flow, respectively, in asthma control assessments. The present study shows that the selection of asthma control criteria among physicians varies and is not always in keeping with current asthma guidelines. Both patients and physicians often consider asthma to be controlled, when according to current guidelines, it is not, and patients consider their asthma better controlled than do physicians. Objective measures of airflow obstruction are rarely used to assess asthma control. The present study stresses the need for improved dissemination - to both patients and physicians - of current recommendations on how asthma control should be determined.


FireProtASR: A Web Server for Fully Automated Ancestral Sequence Reconstruction.
There is a great interest in increasing proteins' stability to widen their usability in numerous biomedical and biotechnological applications. However, native proteins cannot usually withstand the harsh industrial environment, since they are evolved to function under mild conditions. Ancestral sequence reconstruction is a well-established method for deducing the evolutionary history of genes. Besides its applicability to discover the most probable evolutionary ancestors of the modern proteins, ancestral sequence reconstruction has proven to be a useful approach for the design of highly stable proteins. Recently, several computational tools were developed, which make the ancestral reconstruction algorithms accessible to the community, while leaving the most crucial steps of the preparation of the input data on users' side. FireProtASR aims to overcome this obstacle by constructing a fully automated workflow, allowing even the unexperienced users to obtain ancestral sequences based on a sequence query as the only input. FireProtASR is complemented with an interactive, easy-to-use web interface and is freely available at https://loschmidt.chemi.muni.cz/fireprotasr/.


M wave potentiation during and after muscle activity.
The M wave (muscle compound action potential) has been shown to enlarge between successive 3-s voluntary contractions of the human thenar and extensor digitorum brevis (EDB) muscles. The changes, which affected both the amplitude and the area of the M wave, were more obvious in the thenar than in the EDB muscles. In the thenar muscles the mean amplitude was already significantly enlarged after the first voluntary contraction and close to the maximal value by the third (mean maximal increase 23.6 +/- 12.6% of control). The increase in mean M wave area was more gradual, reaching a maximum of 29.3 +/- 14.1% at 100 s. After the voluntary thenar contractions ceased, the amplitude of the M wave subsided more rapidly than the area and had regained the control value within 50 s. The magnitude and time course of the increase in EDB M wave area (maximum change 25.9 +/- 15.2%) were similar to those of the thenar muscles; however, the subsequent decline was slower. The amplitude of the EDB M wave showed the least change, and the maximum increase (11.4 +/- 9.6%) occurred early in the postcontraction period. In both muscles the changes in M wave amplitude and area were significantly different from the control values.


Leukotriene modifier use and asthma severity: how is a new medication being used by adults with asthma?
The introduction of leukotriene modifiers, the first novel class of medications for asthma in more than 2 decades, provided an opportunity to evaluate the clinical context in which patients receive new treatments. Because milder asthma is usually controllable with more familiar medications, we hypothesized that adults with asthma would receive leukotriene modifiers for more severe disease. We conducted a prospective, longitudinal, 18-month cohort study of 349 patients with asthma. We evaluated the association of baseline self-reported medication use and measures of asthma severity. We also examined the impact of baseline measurement of asthma severity on incident leukotriene modifier use at follow-up. At baseline, 39 (11%) of 349 patients reported leukotriene modifier use during the previous 2 weeks (95% confidence interval [CI], 8%-15%). Adults with asthma who reported recent use of leukotriene modifiers were more likely to indicate use of other long-term controller medications for asthma, such as inhaled corticosteroids (80% vs 57%;P =.007). Leukotriene modifier use was also associated with poorer severity-of-asthma scores (mean score difference, 3.6 points; 95% CI, 1.7-5.2 points) and asthma-specific health-related quality of life (mean score difference, 8.1 points; 95% CI, 3.4-12.8 points). Leukotriene modifier users were also more likely to indicate a recent emergency department visit (odds ratio [OR], 2.3; 95% CI, 0.9-5.6) or hospitalization for asthma (OR, 4.1; 95% CI, 1.4-11.4). Greater baseline asthma severity was associated with an increased probability of new-onset leukotriene modifier use during 18-month follow-up. Poorer baseline severity-of-asthma scores and asthma-specific quality-of-life scores were related to a greater likelihood of leukotriene modifier use at follow-up (OR per SD-sized score increment, 2.0; 95% CI, 1.4-2.7; OR, 1.8; 95% CI, 1.3-2.5; respectively). Recent hospitalization for asthma at baseline was also associated with a greater likelihood of leukotriene modifier use at follow-up (OR, 4.9; 95% CI, 1.6-14.8). Adults with asthma who receive leukotriene modifiers have more severe asthma.


Ventilatory constraints during exercise in patients with chronic heart failure.
We examined the degree of ventilatory constraint in patients with a history of chronic heart failure (CHF; n = 11; mean +/- SE age, 62 +/- 4 years; cardiac index [CI], 2.0 +/- 0.1; and ejection fraction [EF], 24 +/- 2%) and in control subjects (CTLS; n = 8; age, 61 +/- 5 years; CI, 2.6 +/- 0.3) by plotting the tidal flow-volume responses to graded exercise in relationship to the maximal flow-volume envelope (MFVL). Inspiratory capacity (IC) maneuvers were performed to follow changes in end-expiratory lung volume (EELV) during exercise, and the degree of expiratory flow limitation was assessed as the percent of the tidal volume (VT) that met or exceeded the expiratory boundary of the MFVL. CHF patients had significantly (p < 0.05) reduced baseline pulmonary function (FVC, 76 +/- 4%; FEV(1), 78 +/- 4% predicted) relative to CTLS (FVC, 99 +/- 4%; FEV(1), 102 +/- 4% predicted). At peak exercise, oxygen consumption (VO(2)) and minute ventilation (V(E)) were lower in CHF patients than in CTLS (VO(2), 17 +/- 2 vs 32 +/- 2 mL/kg/min; VE, 56 +/- 4 vs 82 +/- 6 L/min, respectively), whereas VE/carbon dioxide output was higher (42 +/- 4 vs 29 +/- 5). In CTLS, EELV initially decreased with light exercise, but increased as VE and expiratory flow limitation increased. In contrast, the EELV in patients with CHF remained near residual volume (RV) throughout exercise, despite increasing flow limitation. At peak exercise, IC averaged 91 +/- 3% and 79 +/- 4% (p < 0.05) of the FVC in CHF patients and CTLS, respectively, and flow limitation was present over > 45% of the VT in CHF patients vs < 25% in CTLS (despite the higher VE in CTLS). The least fit and most symptomatic CHF patients demonstrated the lowest EELV, the greatest degree of flow limitation, and a limited response to increased inspired carbon dioxide during exercise, all consistent with VE constraint. We conclude that patients with CHF commonly breathe near RV during exertion and experience expiratory flow limitation. This results in VE constraint and may contribute to exertional intolerance.


Nested communities, invasive species and Holocene extinctions: evaluating the power of a potential conservation tool.
General ecological methods and models that require a minimum amount of information yet are still able to inform conservation planning are particularly valuable. Nested subset analysis has been advocated as such a tool for the prediction of extinction-prone species and populations. However, such advocacy has not been without skepticism and debate, and in the majority of published examples assessing extinction vulnerability, actual extinctions are based on assumptions rather than direct evidence. Here, we empirically test the power of nested subset analysis to predict extinction-prone species, using documented Holocene insular mammal extinctions on three island archipelagos off the west coast of North America. We go on to test whether the introduction of invasive mammals promotes nestedness on islands via extinction. While all three archipelagos were significantly nested before and after the extinction events, nested subset analysis largely failed to predict extinction patterns. We also failed to detect any correlations between the degree of nestedness at the genus-level with area, isolation, or species richness and extinction risk. Biogeography tools, such as nested subset analysis, must be critically evaluated before they are prescribed widely for conservation planning. For these island archipelagos, it appears detailed natural history and taxa-specific ecology may prove critical in predicting patterns of extinction risk.


Inhibition Studies with 2-Bromoethanesulfonate Reveal a Novel Syntrophic Relationship in Anaerobic Oleate Degradation.
Degradation of long-chain fatty acids (LCFAs) in methanogenic environments is a syntrophic process involving the activity of LCFA-degrading bacteria and hydrogen-utilizing methanogens. If methanogens are inhibited, other hydrogen scavengers are needed to achieve complete LCFA degradation. In this work, we developed two different oleate (C18:1 LCFA)-degrading anaerobic enrichment cultures, one methanogenic (ME) and another in which methanogenesis was inhibited (IE). Inhibition of methanogens was attained by adding a solution of 2-bromoethanesulfonate (BrES), which turned out to consist of a mixture of BrES and isethionate. Approximately 5 times faster oleate degradation was accomplished by the IE culture compared with the ME culture. A bacterium closely related to Syntrophomonas zehnderi (99% 16S rRNA gene identity) was the main oleate degrader in both enrichments, in syntrophic relationship with hydrogenotrophic methanogens from the genera Methanobacterium and Methanoculleus (in ME culture) or with a bacterium closely related to Desulfovibrio aminophilus (in IE culture). A Desulfovibrio species was isolated, and its ability to utilize hydrogen was confirmed. This bacterium converted isethionate to acetate and sulfide, with or without hydrogen as electron donor. This bacterium also utilized BrES but only after 3 months of incubation. Our study shows that syntrophic oleate degradation can be coupled to desulfonation.IMPORTANCE In anaerobic treatment of complex wastewater containing fat, oils, and grease, high long-chain fatty acid (LCFA) concentrations may inhibit microbial communities, particularly those of methanogens. Here, we investigated if anaerobic degradation of LCFAs can proceed when methanogens are inhibited and in the absence of typical external electron acceptors, such as nitrate, iron, or sulfate. Inhibition studies were performed with the methanogenic inhibitor 2-bromoethanesulfonate (BrES). We noticed that, after autoclaving, BrES underwent partial hydrolysis and turned out to be a mixture of two sulfonates (BrES and isethionate). We found out that LCFA conversion proceeded faster in the assays where methanogenesis was inhibited, and that it was dependent on the utilization of isethionate. In this study, we report LCFA degradation coupled to desulfonation. Our results also showed that BrES can be utilized by anaerobic bacteria.


Micropuncture measurement of microvascular pressures in isolated lamb lungs during hypoxia.
To study the effect of alveolar hypoxia on the pulmonary circulation of the newborn, we determined the longitudinal distribution of vascular pressures and the distribution of blood flow in isolated blood perfused lungs of newborn lambs during normoxia and hypoxia. We also examined the effect of hypoxia on the slope and intercept of the vascular pressure-flow (P/Q) relation. In 14 lungs, we used the micropipette servonulling technique to measure pressures in 20-80 micron subpleural arterioles and venules during normoxia and hypoxia at constant blood flow. In 5 other lungs, we used radiolabelled microspheres to determine the effect of hypoxia on regional distribution of blood flow. Under baseline normoxic conditions, segmental pressure drops in arteries, microvessels, and veins were approximately equal. During hypoxia pulmonary artery pressure always increased, resulting in a 60% increase in total vascular resistance. All segmental pressure drops increased with hypoxia. Arterial and venous pressure drops increased by 80 and 70% respectively, contributing almost equally to the increase in total pulmonary vascular resistance. Regional distribution of blood flow was uniform during normoxia and was unchanged during hypoxia. Hypoxia increased both the slope and intercept of the P/Q relation significantly, with an increase in fluid filtration rate. We conclude that in the newborn lamb alveolar hypoxia leads to pulmonary arterial and venous constriction, resulting in increased microvascular pressure and fluid filtration.


The nature of the near-infrared features on the venus night side.
Near-infrared images of the Venus night side show bright contrast features that move from east to west, in the direction of the cloud-top atmospheric superrotation. Recently acquired images of the Venus night side along with earlier spectroscopic observations allow identification of the mechanisms that produce these features, their level of formation, and the wind velocities at those levels. The features are detectable only at wavelengths near 1.74 and 2.3 micrometers, in narrow atmospheric windows between the CO(2) and H(2)O bands. The brightest features have brightness temperatures near 480 Kelvin, whereas the darkest features are more than 50 Kelvin cooler. Several factors suggest that this radiation is emitted by hot gases at altitudes below 35 kilometers in the Venus atmosphere. The feature contrasts are produced as this thermal radiation passes through a higher, cooler, atmospheric layer that has horizontal variations in transparency. The 6.5-day east-west rotation period of the features indicates that equatorial wind speeds are near 70 meters per second in this upper layer. Similar wind speeds have been measured by entry probes and balloons at altitudes between 50 and 55 kilometers in the middle cloud layer. The bright features indicate that there are partial clearings in this cloud deck. The presence of these clearings could decrease the efficiency of the atmospheric greenhouse that maintains the high surface temperatures on Venus.


Systemic clearance of p16INK4a -positive senescent cells mitigates age-associated intervertebral disc degeneration.
Age-related changes in the intervertebral discs are the predominant contributors to back pain, a common physical and functional impairment experienced by older persons. Cellular senescence, a process wherein cells undergo growth arrest and chronically secrete numerous inflammatory molecules and proteases, has been reported to cause decline in the health and function of multiple tissues with age. Although senescent cells have been reported to increase in intervertebral degeneration (IDD), it is not known whether they are causative in age-related IDD. The study aimed to elucidate whether a causal relationship exists between cellular senescence and age-related IDD. To examine the impact of senescent cells on age-associated IDD, we used p16-3MR transgenic mice, which enables the selective removal of p16Ink4a -positive senescent cells by the drug ganciclovir. Disc cellularity, aggrecan content and fragmentation alongside expression of inflammatory cytokine (IL-6) and matrix proteases (ADAMTS4 and MMP13) in discs of p16-3MR mice treated with GCV and untreated controls were assessed. In aged mice, reducing the per cent of senescent cells decreased disc aggrecan proteolytic degradation and increased overall proteoglycan matrix content along with improved histological disc features. Additionally, reduction of senescent cells lowered the levels of MMP13, which is purported to promote disc degenerative changes during aging. The findings of this study suggest that systemic reduction in the number of senescent cells ameliorates multiple age-associated changes within the disc tissue. Cellular senescence could therefore serve as a therapeutic target to restore the health of disc tissue that deteriorates with age.


Maternal transmission of multiple sclerosis in a dutch population.
To investigate the parental relationship of patients with multiple sclerosis (MS) from an extended pedigree with extensive genealogical information up to the middle of the 18th century. Multiple sclerosis is a complex disease resulting from genetic and environmental factors. Parent-of-origin effect, a phenomenon when the same allele may express differently depending on the sex of the transmitting parent, may influence the risk for MS. We investigated parental relationships between patients with MS using extensive genealogical information available from the Genetic Research in Isolated Populations program. We compared the average kinship of the parents of MS patients. We further explored the distribution of shortest genealogical links between parents of MS patients. Subjects Twenty-four MS patients from the isolated population who could be linked within a large complex pedigree, including 2471 people in total. The results consistently indicate a higher prevalence of maternal transmission of MS. The kinship between mothers of patients was 3.8 times higher than that between fathers (bootstrap P = .01). Among the 814 shortest connections between parents, 333 were maternal (40.9%, vs 25.0% expected), 98 were paternal (12.0%, vs 25.0% expected), and 383 were maternal-paternal (47.1%, vs 50.0% expected) (P < .001). Mothers of MS patients were more closely related than their fathers. This skewed relationship shows evidence for a maternal effect in MS. The most likely explanation is a gene-environment effect that takes place in utero.


A proposed technique for intraoperative measurement of cervical spine stiffness.
Classically cervical biomechanical data have been obtained using cadaver models. Few methods exist for in vivo measurement of cervical biomechanics. A technique for intraoperative measurement of cervical motion segment stiffness (load-axial displacement) has been developed, enabling load/displacement data to be obtained during surgery at the spinal levels operated on and at adjacent cervical levels. The instrument used for measuring motion segment stiffness was a vertebral retractor equipped with displacement and strain transducers and designed to be placed over the shafts of Caspar distraction screws. Stiffness was measured under simulated anterior cervical surgical conditions. A preliminary study deciphering axial cervical spine load/displacement data on 12 cadavers was performed with this instrument. Using a distraction load of 88.8640 N, axial displacements of 2.110 +/- 0.258 mm (mean +/- standard error of the mean) at C3-C4, 2.367 +/- 0.304 mm at C4-C5, and 2.207 +/- 0.238 mm at C5-C6 were found. When data were evaluated for age, there was a significant decrease in stiffness with advancing age in two of the three spinal segments tested (P < 0.05 for C3-C4, P < 0.04 for C4-C5, P < 0.12 for C5-C6), which correlated with previous studies. These results suggest that this technique may be useful in assessing intraoperative motion-segment stability.


Intrathoracic current flow during transthoracic defibrillation in dogs. Transcardiac current fraction.
To achieve transcardiac threshold current during transthoracic defibrillation, a considerably larger current must be delivered to the thorax to compensate for the shunting effect of the lungs, the thoracic cage, and other elements of the torso. This shunting effect is thus an important determinant of transthoracic defibrillation threshold and can be quantified by the transcardiac current fraction (FC, the ratio of transcardiac to transthoracic threshold currents). Previous estimates of FC have ranged from as low as 3% to as high as 45%. The purpose of of this study was to quantify both FC and the major intrathoracic current pathways. Transthoracic and intrathoracic voltages and currents were simultaneously measured during high-voltage transthoracic shocks in 20 dogs. With correction factors determined from another set of 12 dogs, these raw data were corrected to compensate for field distortion caused by the presence of the intrathoracic electrodes, and the adjusted data were fit to a resistive network model. The results showed that 82% of the transthoracic current was shunted by the thoracic cage, while 14% was shunted by the lungs. The remaining 4% (FC) is the portion that passed through the heart. There was good agreement between the two independent methods used to calculate FC. Analysis based on the model indicated that FC was 3.7%, whereas FC determined by direct measurement with calibrated electrodes was 4.2%. Therefore, the results of this study, in contrast to earlier estimates of FC, show that defibrillation in dogs is achieved by only 4% of the total transthoracic current.


The Prevention Research Centers Healthy Aging Research Network.
The Prevention Research Centers Healthy Aging Research Network (PRC-HAN), funded by the Centers for Disease Control and Prevention's (CDC's) Healthy Aging program, was created in 2001 to help develop partnerships and create a research agenda that promotes healthy aging. The nine universities that participate in the network use their expertise in aging research to collaborate with their communities and other partners to develop and implement health promotion interventions for older adults at the individual, organizational, environmental, and policy levels. The population of older adults in the United States is growing rapidly; approximately 20% of Americans will be aged 65 years or older by 2030. The health and economic impact of an aging society compel the CDC and the public health community to place increased emphasis on preventing unnecessary disease, disability, and injury among older Americans. The PRC-HAN has a broad research agenda that addresses health-promoting skills and behaviors, disease and syndrome topics, and knowledge domains. The network chose physical activity for older adults as its initial focus for research and has initiated two networkwide projects: a comprehensive, multisite survey that collected information on the capacity, content, and accessibility of physical activity programs for older adults and a peer-reviewed publication that describes the role of public health in promoting physical activity among older adults. In addition to participating in the core research area, each network member works independently with its community committee on PRC-HAN activities. As a result, the network is 1) expanding prevention research for older adults and their communities; 2) promoting the translation and dissemination of findings to key stakeholders; 3) strengthening PRC-HAN capacity through partnerships and expanded funding; and 4) stimulating the adoption of policies and programs by engaging policymakers, planners, and practitioners. In 2003, the PRC-HAN initiated an internal evaluation to better define the network's contributions to healthy aging, formalize internal processes, and better equip itself to serve as a model for other PRC thematic networks. The PRC-HAN is conducting a pilot evaluation for eventual inclusion in the PRC national evaluation. The PRC-HAN has established itself as an effective research network to promote healthy aging. It has developed trust and mutual respect among participants, forged strong ties to local communities, and shown the ability to combine its expertise in healthy aging with that of partners in national, state, and local organizations.


Erythropoiesis-stimulating agents in oncology.
Patients who have cancer, particularly those undergoing chemotherapy, frequently become anemic. Therapy with erythropoiesis-stimulating agents (ESAs) is associated with an increase in hemoglobin levels, a reduction in transfusion requirements, and, according to many clinical trialists and experienced clinicians, an improvement in functional status, productivity, and quality of life. Several randomized trials of ESAs in patients who have cancer have recently reported inferior outcomes in tumor progression or survival, raising appropriate concerns about the safety of ESAs in oncology. However, 3 important caveats to these reports exist. First, these clinical trials did not reflect the common use of ESAs in oncology practice (i.e., to treat, rather than prevent, anemia in patients undergoing chemotherapy). Second, the trials were seriously flawed and did not meet reasonable standards for cancer progression or survival trials. Third, during the same period, randomized trials were presented or published that showed no negative impact on tumor progression or survival; these trials have approximately the same shortcomings as trials that suggest a safety issue exists. The lack of definitive answers about the safety of ESAs for treating chemotherapy-related anemia has placed physicians, regulators, and most importantly patients in a difficult position that can only be addressed with additional data. This article reviews relevant preclinical and clinical available data to help improve understanding and guide decision making.


Episodic fluctuations in larval supply
The lack of a clear relationship between spawning output and recruitment success continues to confound attempts to understand and manage temporally variable fish populations. This relationship for a common reef fish is shown to be obscured by nonlinear processes in operation during the larval phase. Nonlinear responses of larval fish to their noisy physical environment may offer a general explanation for the erratic, often episodic, replenishment of open marine populations.


Nitric oxide activates p21ras and leads to the inhibition of endothelial NO synthase by protein nitration.
Recent data has indicated that exogenous nitric oxide (NO) has the ability to decrease endogenous NO production by inhibiting the enzyme responsible for its generation, NO synthase (NOS). Our previous studies have indicated that increased generation of reactive oxygen species (ROS) play an important role in the inhibitory event. However, the mechanisms for these effects remain unclear. Previous studies have suggested that NO can activate p21ras. Thus, the objective of this study was to determine whether NO-mediated activation of p21ras is involved in the inhibitory process, and to further elucidate the involvement of ROS. Using primary cultures of ovine pulmonary arterial endothelial cells we demonstrated that the NO donor SpermineNONOate, increased p21ras activity by 2.3-fold compared to untreated cells, and that the farnesyl-transferase inhibitor, alpha-hydroxyfarnesylphosphonic acid, reduced p21ras activity and significantly reduced inhibition of eNOS. The overexpression of p21ras increased, while the overexpression of an NO unresponsive mutant of p21ras (p21ras C118S) reduced, the inhibition of eNOS by NO. Further, we identified an increase in the level of superoxide and peroxynitrite in endothelial cells exposed to NO that was reduced by p21ras C118S transient transfection. Conversely, levels of superoxide and peroxynitrite could be increased by the over expression of wild type p21ras. Similarly, eNOS nitration induced by NO exposure was reduced by p21ras C118S transient transfection, and increased by the overexpression of wild-type p21ras. Finally, results also demonstrated that eNOS itself was a significant producer of superoxide, and that this appeared to be related to a p21ras-dependent increase in phosphorylation of Ser1177. Our results implicate a signaling pathway involving p21ras activation, superoxide generation, and peroxynitrite formation as being important in the NO-mediated inhibition of eNOS.


Polymer microneedles with interconnected porous structures via a phase inversion route for transdermal medical applications.
Porous polymer microneedles (MNs) have great potential in transdermal medical applications due to their three-dimensional (3D) porous structures and high porosity. However, existing approaches for the fabrication of such porous polymer MNs are complicated and only applicable to limited types of polymers. Here, we describe a facile yet effective phase inversion route to prepare polymer MNs with highly porous and interconnected pore structures. The fabrication process is simple and mild without involving high temperatures or irradiation, and can be applied to a broad spectrum of commonly used polymers (e.g., cellulose acetate (CA), polysulfone (PSF), polyethersulfone (PES), polylactic acid (PLA), etc.). Thanks to the capillary effect and large cavity given by highly porous and interconnected structures, the resulting porous polymer MNs show the capability of rapidly extracting dermal interstitial fluid (ISF) and efficiently loading/releasing drug compounds. As a proof of concept, we demonstrate the use of these porous CA MNs in the highly efficient extraction of ISF for glucose level detection and administration of insulin for hyperglycemia. Given the recent trend of painless techniques in diagnosis and treatment, the current study provides a new opportunity for the fabrication of MN-based devices for transdermal ISF extraction and drug delivery.


[Treatment of refractory malignant biliary obstraction: comparing results of percutaneous transhepatic biliary drainage with external catheters or endoscopic metallic stenting].
To compare the initial results and adverse events of percutaneous transhepatic biliary drainage with external catheters or metallic stents for refractory malignant biliary obstruction. To review the data of those patients with refractory malignant biliary obstruction treated with external catheters or metallic stents in Changhai hospital from Sept. 1999 to Aug. 2002. Liver function tests were determined 2 to 4 days prior to and 5 to 7 days after the procedures and adverse events were recorded. A total of 13 metallic stents were implanted in 13 patients (9 men, 4 women, mean age, 62.5 years) and 16 external catheters were implanted in 15 patients (9 men, 6 women, mean age, 60.8 years). The mean serum total bilirubin concentration prior to and after the procedures were (279.19 +/- 108.15) micro mol/L and (158.02 +/- 99.97) micro mol/L in the metallic stents group, (261.09 +/- 106.48) micro mol/L and (172.81 +/- 106.48) micro mol/L in the external catheters group. The mean serum direct bilirubin concentration were (226.83 +/- 84.03) micro mol/L and (132.57 +/- 80.16) micro mol/L, (208.03 +/- 95.03) micro mol/L and (142.61 +/- 83.74) micro mol/L in the two groups. There is no significant difference of the mean serum total and direct bilirubin concentrations between the two groups prior to the procedures (P > 0.05), but either the mean serum total or direct bilirubin concentration after the procedures in the two groups was significantly lower than that prior to the procedures (P < 0.01). The percentage of changes was significantly greater in the metallic stents group than that in the external catheters group (P < 0.01). Two patients with 3 complications occurred in the metallic stents group, including bile leakage and bleeding (1) and duodenal obstruction (1). Five complications occurred in the external catheters group, including catheter migration (2), catheter occlusion (1), bile leakage (1) and bleeding (1). Percutaneous transhepatic biliary drainage is a reliable and safe therapy for refractory malignant biliary obstruction. The initial results suggest that the potential patency of percutaneous transhepatic biliary drainage with self-expandable metal stent is more rapid and effective than that with external catheters.


Companion animals symposium: microbes and gastrointestinal health of dogs and cats.
Recent molecular studies have revealed complex bacterial, fungal, archaeal, and viral communities in the gastrointestinal tract of dogs and cats. More than 10 bacterial phyla have been identified, with Firmicutes, Bacteroidetes, Proteobacteria, Fusobacteria, and Actinobacteria constituting more than 99% of all gut microbiota. Microbes act as a defending barrier against invading pathogens, aid in digestion, provide nutritional support for enterocytes, and play a crucial role in the development of the immune system. Of significance for gastrointestinal health is their ability to ferment dietary substrates into short-chain fatty acids, predominantly to acetate, propionate, and butyrate. However, microbes can have also a detrimental effect on host health. Specific pathogens (e.g., Salmonella, Campylobacter jejuni, and enterotoxigenic Clostridium perfringens) have been implicated in acute and chronic gastrointestinal disease. Compositional changes in the small intestinal microbiota, potentially leading to changes in intestinal permeability and digestive function, have been suggested in canine small intestinal dysbiosis or antibiotic-responsive diarrhea. There is mounting evidence that microbes play an important role in the pathogenesis of canine and feline inflammatory bowel disease (IBD). Current theories for the development of IBD favor a combination of environmental factors, the intestinal microbiota, and a genetic susceptibility of the host. Recent studies have revealed a genetic susceptibility for defective bacterial clearance in Boxer dogs with granulomatous colitis. Differential expression of pathogen recognition receptors (i.e., Toll-like receptors) were identified in dogs with chronic enteropathies. Similarly to humans, a microbial dysbiosis has been identified in feline and canine IBD. Commonly observed microbial changes are increased Proteobacteria (i.e., Escherichia coli) with concurrent decreases in Firmicutes, especially a reduced diversity in Clostridium clusters XIVa and IV (i.e., Lachnospiraceae, Ruminococcaceae, Faecalibacterium spp.). This would indicate that these bacterial groups, important short-chain fatty acid producers, may play an important role in promoting intestinal health.


Characterization of posttransfusion Plasmodium falciparum infection in semi-immune nonparasitemic patients.
Transfusion-transmitted malaria (TTM) has not been studied with molecular means in hyperendemic areas where it is assumed to occur frequently. The African Investigation of the Mirasol System (AIMS) trial provided the opportunity to study TTM from standard whole blood (WB) units. The Plasmodium genome in transfused WB units and patient samples both before transfusion and 1, 3, 7, and 28 days after transfusion was screened and quantified using real-time polymerase chain reaction. Parasitemic samples were confirmed, three alleles were sequenced, and the percentage homology was determined between paired WB units and patient samples. Anti-Plasmodium titers were quantified by serial dilution. Clinical symptoms and microscopic detection of malaria were monitored. Microscopy was negative below 3 × 10(6) genome copies/mL. Thirty-seven patients who were nonparasitemic before transfusion were exposed to parasitemic WB. The amount of Plasmodium genome load transfused ranged between 0.1 × 10(6) and 2.0 × 10(9) copies. The parasite load received through transfusion by 13 patients with TTM was higher than that received by patients without TTM (p = 0.01). Patients with a single parasitemic posttransfusion sample were not considered to have TTM. Four elements critical to predict outcome emerged: parasite load, patient anti-Plasmodium titer pretransfusion, percentage clearance of parasites at Day 1, and level of anti-Plasmodium humoral immune response. Four patients with TTM became parasite-free at Day 28 (effective control), four patients with TTM maintained relatively stable levels of parasitemia (uncertain control), and five patients reached high levels of parasitemia at Day 28 posttransfusion, indicating ineffective control of malarial infection by semi-immune individuals. TTM in endemic areas is relatively frequent (13 of 112 donations; 11.6%) and, although largely controlled by semi-immunity in recipient patients, may require antimalarial treatment.


Immune checkpoint blockade in malignant mesothelioma.
Malignant mesothelioma (MM) is a highly aggressive malignancy with a very dismal prognosis. Current treatment for unresectable MM is largely unsatisfactory; therefore, new therapeutic approaches are eagerly awaited. A better understanding of the complex mechanisms of immune escape operated by neoplastic cells and the ability to unleash an efficient anti-tumor immune response by targeting regulatory immune checkpoint(s) with immunomodulatory monoclonal antibodies (mAbs), is leading to very promising clinical results in different tumor types. Herein, we highlight the clinical impact so far identified for these new immunomodulatory agents in MM patients and discuss their prospective use to design novel clinical trials.


Development of novel replication-defective lymphocytic choriomeningitis virus vectors expressing SIV antigens.
An important focus in vaccine research is the design of vaccine vectors with low seroprevalence and high immunogenicity. Replication-incompetent lymphocytic choriomeningitis virus (rLCMV) vectors do not elicit vector-neutralizing antibody responses, and homologous prime-boost regimens with rLCMV vectors induce boostable and protective T cell responses to model antigens in mice. However, cellular and humoral immune responses following homologous rLCMV vaccine regimens have not been rigorously evaluated in non-human primates (NHPs). To test whether rLCMV vectors constitute an effective vaccine platform in NHPs, we developed rLCMV vectors expressing SIVmac239 Env and Gag antigens and assessed their immunogenicity in mice and cynomolgus macaques. Immunization with rLCMV vaccine vectors expressing SIV Env and Gag was effective at generating SIV-specific T cell and antibody responses in both mice and NHPs. Epitope mapping using SIV Env in C57BL/6 mice demonstrated that rLCMV vectors induced sustained poly-functional responses to both dominant and subdominant epitopes. Our results suggest the potential of rLCMV vectors as vaccine candidates. Future SIV challenge experiments in rhesus macaques will be needed to assess immune protection by these vaccine vectors.


Repeated administration of cytosine-phosphorothiolated guanine-containing oligonucleotides together with peptide/protein immunization results in enhanced CTL responses with anti-tumor activity.
The development of therapeutic anti-cancer vaccines designed to elicit CTL responses with anti-tumor activity has become a reality thanks to the identification of several tumor-associated Ags and their corresponding peptide T cell epitopes. However, peptide-based vaccines, in general, fail to elicit sufficiently strong CTL responses capable of producing therapeutic anti-tumor effects (i.e., prolongation of survival, tumor reduction). Here we report that repeated administration of synthetic oligonucleotides containing foreign cytosine-phosphorothiolated guanine (CpG) motifs increased 10- to 100-fold the CTL response to immunization with various synthetic peptides corresponding to well-known T cell epitopes. Moreover, repeated CpG administration allowed the induction of CTL to soluble protein even in the absence of additional adjuvant. Our results indicate that the potentiating effect of CpG in CTL responses required the participation of Th lymphocytes. Repeated CpG administration resulted in overt splenomegaly and lymphadenopathy with a significant increase in the numbers of CTL precursors and dendritic cells. Protein vaccination in combination with repeated CpG therapy was effective in delaying tumor cell growth and extending survival in mice bearing melanoma tumors. These findings support the contention that repeated administration of CpG-oligonucleotides enhances the effect of peptide and protein vaccines leading to potent anti-tumor responses, presumably through the induction of Th1 and dendritic cells, which are essential for optimal CTL responses. The immunostimulatory properties of CpG motifs may be key in inducing a consistent long term immunity to tumor-associated Ags when using peptides or proteins as T cell-inducing vaccines.


It takes patience and persistence to get negative feedback about patients' experiences: a secondary analysis of national inpatient survey data.
Patient experience surveys are increasingly used to gain information about the quality of healthcare. This paper investigates whether patients who respond before and after reminders to a large national survey of inpatient experience differ in systematic ways in how they evaluate the care they received. The English national inpatient survey of 2009 obtained data from just under 70,000 patients. We used ordinal logistic regression to analyse their evaluations of the quality of their care in relation to whether or not they had received a reminder before they responded. 33% of patients responded after the first questionnaire, a further 9% after the first reminder, and a further 10% after the second reminder. Evaluations were less positive among people who responded only after a reminder and lower still among those who needed a second reminder. Quality improvement efforts depend on having accurate data and negative evaluations of care received in healthcare settings are particularly valuable. This study shows that there is a relationship between the time taken to respond and patients' evaluations of the care they received, with early responders being more likely to give positive evaluations. This suggests that bias towards positive evaluations could be introduced if the time allowed for patients to respond is truncated or if reminders are omitted.


The frequency and extent of mammillary body atrophy associated with surgical removal of a colloid cyst.
Patients who have had a colloid cyst removed from the third ventricle sometimes experience some difficulty with day-to-day memory. This study provided quantitative MR imaging volume measures of 1 structure potentially responsible for mnemonic problems, the mammillary bodies. Additional volume estimates in structures connected to the mammillary bodies sought to determine the specificity of any atrophy. Volume estimates of the mammillary bodies were performed on 38 patients after surgical removal of colloid cysts and 20 control subjects by the application of stereologic volume-estimation techniques. For the mammillary body measures, 2 groups of MR images were assessed (0.8- and 1.0-mm section thickness) to compare the sensitivity of each imaging sequence for detecting any atrophy. Other structures associated with memory processes, such as the hippocampus and fornix, were also assessed quantitatively to determine whether there was a correlation between mammillary body damage and atrophy in connecting structures. Our investigations established the superiority of 0.8-mm-volume scans over standard isotropic 1.0-mm-thick-volume scans for mammillary body assessments. Comparisons with 20 age-matched controls revealed that patients with colloid cysts frequently showed significant mammillary body atrophy (mean volume of colloid cysts, 0.037 cm(3) right and 0.038 cm(3) left; control subjects, 0.069 cm(3) right and 0.067 cm(3) left). In fact, every patient had a mammillary body volume below the control mean, and the majority of patients had a volume decrease of >1 SD (82% right, 74% left). Mammillary body volumes correlated with fornix volumes in the same patient group. Our results reveal the frequent presence of mammillary body atrophy in patients with surgical removal of colloid cysts and indicate that this atrophy is partly due to a loss of temporal lobe projections in the fornix.